There has been significant progress over the last 15 years in the development of both methods and systems that provide real-time detection and classification of airborne biological and non-biological particles. Most of the methods and systems have been based on optical approaches that utilize both particle sizing and laser induced fluorescence (LIF) to discriminate biological from non-biological particles. These methods have included illumination approaches using various wavelengths specific towards exciting endogenous fluorophores commonly found in biological particles. LIF based approaches have also included multiple excitation sources operating at different wavelengths for improved classification. Other approaches have included laser breakdown spectroscopy coupled with LIF detection for measuring both auto-fluorescence and mineral content of single particles, as well as, Raman spectroscopy and Fourier transform infrared spectroscopy of single particles.
There is a growing need for improved environmental biosurveillance in such areas as battlefield and homeland defense, indoor and outdoor air quality monitoring, airborne hospital infection control, contamination control monitoring of biopharmaceutical and other manufacturing operations, sewage plants, animal production houses and other operations where continuous real-time monitoring helps provide early warning and prevention of harmful exposure of microorganisms, viruses and other types of biological particles. With respect to defense applications, the deadliest form of a biological attack is from aerosolized agents. To date, optical methods that utilize both particle sizing and laser induced fluorescence have been applied to battlefield and homeland defense. This approach has proven to be an effective early warning capability, in particular, for building protection applications that monitor for indoor biological attacks. To a lesser extent, real-time airborne microbial monitoring using LIF based detection has been applied towards contamination control monitoring of biopharmaceutical manufacturing and other clean room operations. While these LIF based biological particle counter methods have shown to be useful early warning systems, they are not without limitations and significant room for improvement exists.
There are three primary limitations with LIF based biological particulate detection. First, is their ability to detect single vegetative or spore type organisms. Current fielded LIF based biological particle detectors are limited to detection of vegetative cell and spore aggregate particles or particles that contain numerous vegetative cells or spores per aerosol particle. For both biodefense and other applications the detection of single vegetative cells and spores is required and necessary to provide adequate protection or effective contamination control monitoring. For LIF based approaches, unless a laser source with considerable optical power is used an insufficient amount of light is emitted from fluorescence excitation to reliably and accurately classify a biological from a non-biological particle when the particle size is in the 0.5-1.5 micron diameter in size. This is particularly true for longer excitation wavelengths such as the 350-450 nm wavelength range but also applies, in most instances, for shorter wavelengths such as the 250-300 nm wavelength range.
Second, their ability to discriminate biological from non-biological particles and to classify biological particle types, such as bacterial spores, mold spores, vegetative bacteria, viral aggregates, protein toxin aggregates, and fomite particles containing bacteria, viruses, or fungi is very limited. With a minimum need for discrimination of biological from non-biological, the use of LIF based particle detection approaches are useful for discrimination from inorganic particles and non-fluorescing man-made particles but face serious limitations for particles that have been doped with fluorophores, such as paper particles or clothing particles containing optical brighteners, and commonly encountered particles that have intrinsic fluorescence, such as human skin cell fragments and animal dander. The use of LIF based multi-wavelength excitation approaches have some improved classification over single source approaches but the approaches are not cost effective for widespread application.
Third, current LIF based approaches are limited in their ability to detect biological particle concentration levels of interest, particularly in applications where detection of only a fraction to a few particles per liter of air is required. The ability to both detect in a timely manner and to discriminate these low levels from commonly encountered aerosol backgrounds becomes very challenging for LIF based biological particle detection approaches. Current systems are limited to 1-3 liter per minute air sampling flow rates. This equates to slow response times and long sampling times in order to detect and alarm on the presence of a low concentration of biological particles. The use of aerosol concentrators to compensate for this are limited in their application because the problem of discrimination increases with the increase in sampling flow rate and background aerosol concentration.